Adenoviruses are eukaryotic DNA viruses that can be modified to efficiently deliver a therapeutic or reporter transgene to a variety of cell types. Human adenoviruses are comprised of a linear, approximately 36 kb double-stranded DNA genome, which is divided into 100 map units (m.u.), each of which is 360 bp in length. The DNA contains short inverted terminal repeats (ITR) at each end of the genome that are required for viral DNA replication. The gene products are organized into early (E1 through E4) and late (L1 through L5) regions, based on expression before or after the initiation of viral DNA synthesis [see, e.g., M. S. Horwitz et al, "Adenoviridae and Their Replication", Virology, second edition, pp. 1712, ed. B. N. Fields et al, Raven Press Ltd., New York (1990)]. The adenoviruses types 2 and 5 (Ad2 and Ad5, respectively), are not associated with human malignancies.
Recombinant adenoviruses are capable of providing extremely high levels of transgene delivery to virtually all cell types, regardless of the mitotic state. The efficacy of this system in delivering a therapeutic transgene in vivo that complements a genetic imbalance has been demonstrated in animal models of various disorders [K. F. Kozarsky et al, Somatic Cell Mol. Genet., 19:449-458 (1993) ("Kozarsky et al I"); K. F. Kozarsky et al, J. Biol. Chem., 269:13695-13702 (1994) ("Kozarsky et al II); Y. Watanabe, Atherosclerosis, 36:261-268 (1986); K. Tanzawa et al, FEBS Letters, 118(1):81-84 (1980); J. L. Golasten et al, New Engl. J. Med., 309(11983):288-296 (1983); S. Ishibashi et al, J. Clin. Invest., 92:883-893 (1993); and S. Ishibashi et al, J. Clin. Invest., 93:1885-1893 (1994)]. The use of recombinant adenoviruses in the transduction of genes into hepatocytes in vivo has previously been demonstrated in rodents and rabbits [see, e.g., Kozarsky II, cited above, and S. Ishibashi et al, J. Clin. Invest., 92:883-893 (1993)].
The first-generation recombinant, replication-deficient adenoviruses which have been developed for gene therapy contain deletions of the entire E1a and part of the E1b regions. This replication-defective virus is grown on an adenovirus-transformed, complementation human embryonic kidney cell line containing a functional adenovirus E1a gene which provides a transacting E1a protein, the 293 cell [ATCC CRL1573]. E1-deleted viruses are capable of replicating and producing infectious virus in the 293 cells, which provide E1a and E1b region gene products in trans. The resulting virus is capable of infecting many cell types and can express the introduced gene (providing it carries its own promoter), but cannot replicate in a cell that does not carry the E1 region DNA unless the cell is infected at a very high multiplicity of infection.
However, in vivo studies revealed transgene expression in these E1 deleted vectors was transient and invariably associated with the development of severe inflammation at the site of vector targeting [S. Ishibashi et al, J. Clin. Invest., 93:1885-1893 (1994); J. M. Wilson et al, Proc. Natl. Acad. Sci., USA, 85:4421-4424 (1988); J. M. Wilson et al, Clin. Bio., 3:21-26 (1991); M. Grossman et al, Som. Cell. and Mol. Gen., 17:601-607 (1991)]. Antigenic targets for immune mediated clearance are viral proteins expressed from the recombinant viral genome and/or the product of the transgene [Y. Yang et al, Proc. Natl. Acad. Sci., USA, 91:4407-4411 (May 1994); Y. Yang et al, Immun., 1:433-442 (August 1994)].
There remains a need in the art for additional recombinant adenoviruses, therapeutic compositions and methods which enable effective treatment of disorders and diseases by gene therapy.